PROJECT SUMMARY Hematopoietic stem cells (HSCs) are undifferentiated, long-lived cells that give rise to lineage-specific progenitors and retain their stem cell identity by undergoing self-renewal. HSCs have been widely used in transplantation to treat patients with hematologic cancers, solid tumors and autoimmune diseases, and are also promising cell targets for gene therapies that can potentially treat a broad variety of human diseases. However, development of these important clinical applications of HSCs is severely hampered by the difficulty in expanding these cells due to the lack of understanding of the intracellular and extracellular signals that govern HSC fates for repopulation, differentiation and quiescence. Yin Yang (YY1) is a ubiquitous transcription factor and mammalian Polycomb Group Protein (PcG) with important functions that regulate embryonic development, lineage differentiation, and cell proliferation. Our studies showed that YY1 is a unique stem cell promoting factor as high YY1 expression plays a double-edged sword by expanding HSCs while decreasing B cells in vivo. Thus, targeting YY1 may lead into great therapeutic potential by increasing bone marrow transplant efficiency and by decreasing transplant rejection. Our studies showed that YY1 occupies promoter and enhancer sequences and activates Kit, the gene encoding for the key receptor tyrosine kinase for HSC quiescence. YY1 is a critical regulator of cohesin expression in hematopoietic stem progenitor cells, and YY1 plays a critical role in HSC self-renewal and differentiation. Herein, we will further elucidate the molecular mechanisms by which YY1 promotes HSC self-renewal. We will determine whether YY1 promotes HSC activities through control of the Kit locus, and/or through its regulation of the key chromosome structural maintenance complex, cohesin. The work proposed here is a direct extension of work carried out in my SERCA K01 studies. The proposal will provide foundational data on mechanisms governing HSC function and may lead to paradigm-shifting models on the general, as well as specific control of chromatin structure by YY1. These studies will generate key mouse models and molecular insights to drive a future hypothesis-driven R01 application focusing on the control of high-order chromosome structure in HSC biology and the impact of YY1 regulation of cohesin in malignant hematopoiesis. Given the need to develop more effective strategies to expand HSCs in therapeutic contexts, elucidating the molecular mechanisms of YY1 regulation of HSCs will be highly significant.